Protein kinase c inhibitors for treatment of uveal melanoma

ABSTRACT

The present disclosure relates to use of a protein kinase C (PKC) inhibitor in the treatment or prevention of proliferative diseases. The disclosure also relates to corresponding pharmaceutical formulations, uses, methods, combinations, and related disclosure embodiments. The disclosure further relates to use of a pharmaceutical combination comprising a PKC inhibitor and another therapeutic agent, such as an MDM2 inhibitor, in the treatment or prevention of a proliferative disease.

RELATED APPLICATIONS

This application claims priority to, and the benefit of, U.S.Application Ser. No. 62/557,520, filed Sep. 12, 2017, the content ofwhich is hereby incorporated by reference in its entirety.

BACKGROUND

Uveal melanoma (UM) is the most common cancer of the eye in adults(Singh A D. et al., Ophthalmology. 2011; 118: 1881-5). Most UM patientsdevelop metastases for which no curative treatment has been identifiedso far.

The majority of UM tumors have mutations in the genes GNAQ (guaninenucleotide-binding protein G(q) subunit alpha) and GNA11 (guaninenucleotide-binding protein G(q) subunit 11), which encode for smallGTPases (Harbour J W. Pigment Cell Melanoma Res. 2012; 25:171-81). Bothof these mutations lead to activation of the protein kinase C (PKC)pathway. The up-regulation of PKC pathway has downstream effects whichleads to constitutive activation of the mitogen-activated protein kinase(MAPK) signaling pathway that has been implicated in causinguncontrolled cell growth in a number of proliferative diseases.

Whilst anti-proliferative effects have been observed with certain PKCpathway inhibitors, no sustained MAPK pathway inhibition has beenobserved. Thus far, PKC inhibitors (PKCi) have had limited efficacy assingle agents in patients (Mochly-Rosen D et al., Nat Rev Drug Discov.2012 December; 11(12):937-57). There still remains an unmet need toprovide next generation PKC inhibitors for treating uveal melanoma.

SUMMARY

In one aspect, the present disclosure features a method for thetreatment or prevention of a proliferative disease, e.g., uvealmelanoma, in a subject in need thereof. The method includesadministration of a therapeutically effective amount of a protein kinaseC inhibitor (PKCi) in said subject.

The method can include one or more of the following features.

In certain embodiments, the PKC inhibitor is3-amino-N-(3-(4-amino-4-methylpiperidin-1-yl)pyridin-2-yl)-6-(3-(trifluoromethyl)pyridin-2-yl)pyrazine-2-carboxamide(Compound I) or a pharmaceutically acceptable salt thereof.

In certain embodiments, the PKC inhibitor is administered orally.

In certain embodiments, the PKC inhibitor is administered on an emptystomach, such as at least 1 hour before or 2 hours after a meal. Forexample, water and other medications may be permitted during thisperiod. In other embodiments, the PKC inhibitor is administeredimmediately prior to or after the subject eats some food.

In certain embodiments, the method includes administering to a subjectin need of thereof3-amino-N-(3-(4-amino-4-methylpiperidin-1-yl)pyridin-2-yl)-6-(3-(trifluoromethyl)pyridin-2-yl)pyrazine-2-carboxamide(Compound I) or a pharmaceutically acceptable salt thereof at a dose offrom about 100 mg to about 1000 mg of Compound I daily.

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered at a dose of between about 50 mg twice a day(BID) and about 400 mg BID of Compound I (e.g., at a dose of about 50 mgBID, 100 mg BID, 150 mg BID, 200 mg BID, 250 mg BID, 300 mg BID, 350 mgBID, or 400 mg BID).

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered at a dose of about 300 mg BID of Compound I.

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered at a dose of from about 100 mg to about 1000 mgof Compound I once daily (e.g., about 400 mg, 500 mg, 600 mg, 700 mg orabout 800 mg once daily).

In certain embodiments, the subject suffers from metastatic uvealmelanoma.

In certain embodiments, the subject is a human being, e.g., an adultpatient aged 18 years or older.

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is orally administered.

In certain embodiments, the method further includes administering to thesubject one or more other therapeutic agents, e.g., a mouse doubleminute 2 inhibitor (MDM2i or HDM2i). MDM2 and HDM2, the human isoform ofMDM2, are used interchangeably herein.

In certain embodiments, the method further includes administering to thesubject a mouse double minute 2 inhibitor (MDM2i) selected from(S)-5-(5-chloro-1-methyl-2-oxo-1,2-dihydro-pyridin-3-yl)-6-(4-chloro-phenyl)-2-(2,4-dimethoxy-pyrimidin-5-yl)-1-isopropyl-5,6-dihydro-1H-pyrrolo[3,4-d]imidazol-4-one(Compound II),(S)-1-(4-chloro-phenyl)-7-isopropoxy-6-methoxy-2-(4-{methyl-[4-(4-methyl-3-oxo-piperazin-1-yl)-trans-cyclohexylmethyl]-amino}-phenyl)-1,4-dihydro-2H-isoquinolin-3-one(Compound III), and pharmaceutically acceptable salts thereof.

In certain embodiments, the method further includes administering to thesubject(S)-1-(4-chloro-phenyl)-7-isopropoxy-6-methoxy-2-(4-{methyl-[4-(4-methyl-3-oxo-piperazin-1-yl)-trans-cyclohexylmethyl]-amino}-phenyl)-1,4-dihydro-2H-isoquinolin-3-one(Compound III), or a pharmaceutically acceptable salt thereof, forexample, orally.

In certain embodiments, the method further includes administering to thesubject(S)-5-(5-chloro-1-methyl-2-oxo-1,2-dihydro-pyridin-3-yl)-6-(4-chloro-phenyl)-2-(2,4-dimethoxy-pyrimidin-5-yl)-1-isopropyl-5,6-dihydro-1H-pyrrolo[3,4-d]imidazol-4-one(Compound II), or a pharmaceutically acceptable salt thereof, forexample, orally.

In certain embodiments, the method further includes administering to thesubject(S)-5-(5-chloro-1-methyl-2-oxo-1,2-dihydro-pyridin-3-yl)-6-(4-chloro-phenyl)-2-(2,4-dimethoxy-pyrimidin-5-yl)-1-isopropyl-5,6-dihydro-1H-pyrrolo[3,4-d]imidazol-4-one(Compound II), or a pharmaceutically acceptable salt thereof onceweekly, for example, orally.

In certain embodiments, Compound II or a pharmaceutically acceptablesalt thereof is administered at a dose of from about 50 mg to about 200mg of Compound II (e.g., at a dose of about 50 mg, about 80 mg, about100 mg, about 120 mg, about 150 mg, or about 200 mg of Compound II).

In certain embodiments, Compound II or III, or a pharmaceuticallyacceptable salt thereof is orally administered.

In certain embodiments, Compound II or III, or a pharmaceuticallyacceptable salt thereof, is administered on an empty stomach, such as atleast 1 hour before or 2 hours after a meal. For example, water andother medications may be permitted during this period. In otherembodiments, Compound II or III, or a pharmaceutically acceptable saltthereof, is administered immediately prior to or after the subject eatssome food.

In certain embodiments, the uveal melanoma or metastatic uveal melanomacomprises functional p53 or wild-type TP53.

In certain embodiments, the uveal melanoma or metastatic uveal melanomais characterized by mutation of guanine nucleotide-binding protein G(q)subunit alpha (GNAQ) gene or guanine nucleotide-binding protein G(q)subunit 11 (GNA11) gene.

The following disclosure pertains to dually targeting p53, either aloneor in combination with the PKC pathway in order to treat UM. In thisway, the MDM2 inhibitor promotes the beneficial effect of anothercompound that targets a possibly subordinate, interdependent or simplycoexisting biochemical pathway implicated in causing a proliferativedisease.

In another aspect, the present disclosure features a PKC inhibitor foruse in treating or preventing a proliferative disease, e.g., uvealmelanoma, either alone or in a combination therapy with one or moreother therapeutic agents such as an MDM2 inhibitor. For example, the PKCinhibitor is Compound I or a pharmaceutically acceptable salt thereof.For example, the PKC inhibitor is suitable for administration at a doseof from about 100 mg to about 1000 mg of Compound I daily, e.g., viaoral administration. For example, the PKC inhibitor is administered onan empty stomach, such as at least 1 hour before or 2 hours after ameal.

In yet another aspect, the present disclosure features use of a PKCinhibitor in the manufacture of a medicament for the treatment orprevention of a proliferative disease, e.g., uveal melanoma, eitheralone or in a combination therapy with one or more other therapeuticagents such as an MDM2 inhibitor. For example, the PKC inhibitor isCompound I or a pharmaceutically acceptable salt thereof. For example,the PKC inhibitor is suitable for administration at a dose of from about100 mg to about 1000 mg of Compound I daily, e.g., via oraladministration. For example, the PKC inhibitor is administered on anempty stomach, such as at least 1 hour before or 2 hours after a meal.

Any of the above aspects and embodiments can be combined with any otheraspect or embodiment.

Unless otherwise defined, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. In the specification, thesingular forms also include the plural unless the context clearlydictates otherwise. Although methods and materials similar or equivalentto those described herein can be used in the practice or testing of thepresent invention, suitable methods and materials are described below.All publications, patent applications, patents and other referencesmentioned herein are incorporated by reference. The references citedherein are not admitted to be prior art to the claimed invention. In thecase of conflict, the present specification, including definitions, willcontrol. In addition, the materials, methods and examples areillustrative only and are not intended to be limiting.

Other features and advantage of the disclosure will be apparent from thefollowing detailed description and claims.

DETAILED DESCRIPTION OF THE DISCLOSURE

Protein kinase C, specifically isoforms alpha and/or theta (PKCα/θ), hasbeen found to play a role in certain disorders in a human or animalsubject. The present disclosure relates to compounds, e.g., PKCinhibitors, that exhibit anti-proliferative activity when used alone andin combination, preferably in uveal melanoma (UM) patients. Suitably,the method relates to methods of treating a proliferative disease byadministration or co-administration of said compounds.

In some embodiments, the PKC inhibitor as used herein can be3-amino-N-(3-(4-amino-4-methylpiperidin-1-yl)pyridin-2-yl)-6-(3-(trifluoromethyl)pyridin-2-yl)pyrazine-2-carboxamide(Compound I) of formula I:

(also known as LXS196). In some embodiments, the PKC inhibitor as usedherein can be a pharmaceutically acceptable salt of Compound I.

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered, e.g., orally administered at a dose of fromabout 100 mg to about 1000 mg of Compound I daily (e.g., about 100-800mg or about 100-400 mg daily). For example, Compound I or apharmaceutically acceptable salt thereof is administered at a dose ofbetween about 50 mg twice a day (BID) and about 400 mg BID of Compound I(e.g., at a dose of about 50 mg BID, 100 mg BID, 150 mg BID, 200 mg BID,250 mg BID, 300 mg BID, 350 mg BID, or 400 mg BID). As another example,Compound I or a pharmaceutically acceptable salt thereof is administeredat a dose of from about 100 mg to about 1000 mg of Compound I once daily(e.g., about 400 mg, 500 mg, 600 mg, 700 mg or about 800 mg once daily).

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered, e.g., orally administered at a dose of fromabout 1 mg/kg to about 25 mg/kg (e.g., between about 2 mg/kg to about 20mg/kg, or between about 3 mg/kg to about 15 mg/kg, or between about 4mg/kg to about 10 mg/kg) of Compound I daily based on the body weight ofthe subject. For example, Compound I or a pharmaceutically acceptablesalt thereof is administered at a dose of between about 1 mg/kg twice aday (BID) and about 8 mg/kg BID of Compound I (e.g., at a dose of about1 mg/kg BID, 2 mg/kg BID, 3 mg/kg BID, 4 mg/kg BID, 5 mg/kg BID, 6 mg/kgBID, 7 mg/kg BID, or 8 mg/kg BID). As another example, Compound I or apharmaceutically acceptable salt thereof is administered at a dose offrom about 2 mg/kg to about 20 mg/kg of Compound I once daily (e.g.,about 3 mg/kg, 5 mg/kg, 8 mg/kg, 10 mg/kg, 12 mg/kg, 15 mg/kg, or 18mg/kg once daily).

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered at a dose that is therapeutically efficientwhile having little or no adverse effect. For example, said dose causeslittle or no hypotension. For example, about 5% or less (e.g., about2.5% or less, 2% or less, 1% or less, or about 0.5% or less) patientsadministered with said dose experience hypotension of grade 3 or 4.

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered on an empty stomach, such as at least 1 hourbefore or 2 hours after a meal. For example, water and other medicationsmay be permitted during this period.

In other embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered immediately prior to or after the subject eatssome food.

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered alone.

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered with one or more other therapeutic agents, e.g.,an anti-cancer agent.

In certain embodiments, Compound I or a pharmaceutically acceptable saltthereof is administered (either alone or in combination with anothertherapeutic agent) for a period of time (e.g., 1, 2, 3, 4, 5, 6, 7, 8,9, 10, 11, 12, 13, or 14 days, specifically for example, administereddaily for 7 days) at a lower dose immediately prior to treatment of apharmaceutical combination comprising Compound I or a pharmaceuticallyacceptable salt thereof and one or more other therapeutic agents, e.g.,an anti-cancer agent.

The present disclosure, relates also to a pharmaceutical combination,especially a pharmaceutical combination product, comprising, e.g.,Compound I or a pharmaceutically acceptable salt thereof and one or moreof other therapeutic agents described herein, and at least onepharmaceutically acceptable carrier.

In certain embodiments, the uveal melanoma or metastatic uveal melanomacomprises functional p53 or wild-type TP53. In other embodiments, theuveal melanoma or metastatic uveal melanoma comprises mutated TP53.

The protein p53 is a transcription factor that controls the expressionof a multitude of target genes involved in DNA damage repair, apoptosisand cell cycle arrest, which are all important phenomena counteractingthe malignant growth of tumors. The TP53 gene is one of the mostfrequently mutated genes in human cancers, with approximately half ofall cancers having inactivated p53. Furthermore, in cancers with anon-mutated TP53 gene, typically the p53 is functionally inactivated atthe protein level. One of the mechanisms of p53 inactivation is throughits interaction with human homolog of MDM2 (Mouse double minute 2)protein. MDM2 protein functions both as an E3 ubiquitin ligase, thatleads to proteasomal degradation of p53, and an inhibitor of p53transcriptional activation. Therefore, MDM2 is an important negativeregulator of the p53 tumor suppressor. MDM2 inhibitors can preventinteraction between MDM2 and p53 and thus allow the p53 protein to exertits effector functions. Whilst TP53 mutations are not common in UM,there are reports suggesting the p53 pathway is inactivated by eitherhigh expression of MDM2 protein or downregulation of the PERP protein inUM patients (Davies et al., Cell Death Dis. 2011 March; 2(3): e136).

A combination of an MDM2 inhibitor (Nutlin-3) has been shown to actsynergistically with reactivation of p53 and induction of tumor cellapoptosis (RITA) and Topotecan to cause growth inhibition in UM celllines (De Lange J. et al., Oncogene. 2012; 31:1105-16). However,Nutlin-3 and Topotecan delayed in vivo tumor growth only in a limitedmanner.

In some embodiments, the PKC inhibitor as used herein can be used in acombination therapy with one or more other therapeutic agents, such as amouse double minute 2 inhibitor (MDM2i or HDM2i). By targeting the PKCpathway, either alone or in combination with p53, the PKC inhibitorsdisclosed herein or pharmaceutical compositions thereof andpharmaceutical combinations provided herein have been surprisingly foundto be useful in treating UM or metastatic UM. The pharmaceuticalcompositions and pharmaceutical combinations and/or drug regimensdescribed herein led to the induction of cell death in vitro, tumorstabilization and even tumor regression in vivo, with a surprisinglyhigh in vivo tumor shrinkage observed in one combination.

The MDM2 inhibitor suitable for the methods, pharmaceutical compositionsand/or pharmaceutical combinations described herein can be(S)-5-(5-chloro-1-methyl-2-oxo-1,2-dihydro-pyridin-3-yl)-6-(4-chloro-phenyl)-2-(2,4-dimethoxy-pyrimidin-5-yl)-1-isopropyl-5,6-dihydro-1H-pyrrolo[3,4-d]imidazol-4-one(Compound II) of formula II:

The compound of formula II can be prepared as described inWO2013/111105.

The MDM2 inhibitor suitable for the methods, pharmaceutical compositionsand/or pharmaceutical combinations described herein can also be(S)-1-(4-chloro-phenyl)-7-isopropoxy-6-methoxy-2-(4-{methyl-[4-(4-methyl-3-oxo-piperazin-1-yl)-trans-cyclohexylmethyl]-amino}-phenyl)-1,4-dihydro-2H-isoquinolin-3-one(compound III) of formula III:

Compound III can be prepared as described in WO2011/076786. In oneembodiment, the the pharmaceutically acceptable salt of Compound III isa bisulphate salt. Crystalline form of the bisulfate salt of CompoundIII is described in WO2012/066095.

Other MDM2 inhibitors suitable for the methods, pharmaceuticalcompositions and/or pharmaceutical combinations described herein canalso be those known by a skilled artisan, such as peptide MDM2inhibitors, chlorofusin, spiro-oxindoles, Nutlins, benzodiazepinediones,NU8231, NU8354, NSC-66811, orN¹-(2-(1H-indol-3-yl)ethyl)-N⁴-(pyridin-4-yl)benzene-1,4-diamine (see,e.g., Sharmila Patel & Mark R Player (2008) Small-molecule inhibitors ofthe p53-HDM2 interaction for the treatment of cancer, Expert Opinion onInvestigational Drugs, 17:12, 1865-1882, which is incorporated herein inits entirety).

The present disclosure encompasses embodiments that include allpharmaceutically acceptable salts of the compounds useful according tothe disclosure provided herein. As used herein, “pharmaceuticallyacceptable salt” refers to derivatives of the disclosed compoundswherein the parent compound is modified by converting an existing acidor base moiety to its salt form. Examples of pharmaceutically acceptablesalts include, but are not limited to, mineral or organic acid salts ofbasic residues such as amines; alkali or organic salts of acidicresidues such as carboxylic acids; and the like. The pharmaceuticallyacceptable salts of the present disclosure include the conventionalnon-toxic salts of the parent compound formed, for example, fromnon-toxic inorganic or organic acids. The pharmaceutically acceptablesalts of the present disclosure can be synthesized from the parentcompound which contains a basic or acidic moiety by conventionalchemical methods. Generally, such salts can be prepared by reacting thefree acid or base forms of these compounds with a stoichiometric amountof the appropriate base or acid in water or in an organic solvent, or ina mixture of the two; generally, non-aqueous media like ether, ethylacetate, ethanol, isopropanol, or acetonitrile are preferred. Lists ofsuitable salts are found in Remington's Pharmaceutical Sciences, 17^(th)ed., Mack Publishing Company, Easton, Pa., 1985, p. 1418 and Journal ofPharmaceutical Science, 66, 2 (1977), each of which is incorporatedherein by reference in its entirety. For example, preferredpharmaceutically acceptable salts include, but are not limited to,mineral or organic acid salts of basic residues such as amines. Forexample, the salt can be a sulphate salt, or bisulphate salt.

The compounds disclosed herein, particularly Compound I, may be in aform of a pharmaceutically acceptable prodrug. The term“pharmaceutically acceptable prodrugs” as used herein refers to thoseprodrugs of the compounds of the present disclosure which are, withinthe scope of sound medical judgment, suitable for use in contact withthe tissues of humans and lower animals without undue toxicity,irritation, allergic response, and the like, commensurate with areasonable benefit/risk ratio, and effective for their intended use, aswell as the zwitterionic forms, where possible, of the compounds of thedisclosure. The term “prodrug” refers to compounds that are rapidlytransformed in vivo to yield the parent compound of the above formula,for example by hydrolysis in blood. A thorough discussion is provided inT. Higuchi and V. Stella, Pro-drugs as Novel Delivery Systems, Vol. 14of the A.C.S. Symposium Series, and in Edward B. Roche, ed.,Bioreversible Carriers in Drug Design, American PharmaceuticalAssociation and Pergamon Press, 1987, both of which are incorporatedherein by reference.

The phrase “pharmaceutically acceptable” as employed herein refers tothose compounds, materials, compositions, and/or dosage forms which are,within the scope of sound medical judgment, suitable for use in contactwith the tissues of human beings and animals without excessive toxicity,irritation, allergic response, or other problem or complication,commensurate with a reasonable benefit/risk ratio.

Any formula given herein is also intended to represent unlabeled formsas well as isotopically labeled forms of the compounds. Isotopicallylabeled compounds have structures depicted by the formulae given hereinexcept that one or more atoms are replaced by an atom having a selectedatomic mass or mass number. Isotopes that can be incorporated intocompounds of the disclosure include, for example, isotopes of hydrogen,carbon, nitrogen, oxygen, fluorine, and chlorine, such as ³H, ¹¹C, ¹³C,¹⁴C, ¹⁵N, ¹⁸F, and ³⁶Cl. Accordingly, it should be understood that thepresent disclosure includes compounds that incorporate one or more ofany of the aforementioned isotopes, including for example, radioactiveisotopes, such as ³H and ¹⁴C, or those into which non-radioactiveisotopes, such as ²H and ¹³C are present. Such isotopically labelledcompounds are useful in metabolic studies (with ¹⁴C), reaction kineticstudies (with, for example ²H or ³H), detection or imaging techniques,such as positron emission tomography (PET) or single-photon emissioncomputed tomography (SPECT) including drug or substrate tissuedistribution assays, or in radioactive treatment of patients. Inparticular, an ¹⁸F or labeled compound may be particularly desirable forPET or SPECT studies. Isotopically-labeled compounds can generally beprepared by conventional techniques known to those skilled in the art,e.g., using an appropriate isotopically-labeled reagents in place of thenon-labeled reagent previously employed.

The compounds described herein, e.g., a PKC inhibitor and/or a MDM2inhibitor, can be used in combination, especially for use in apharmaceutical combination that may optionally include further co-agentsas defined below. All of these materials may be referred to as “activeingredients” in the combination. It should be understood that both terms(e.g., compound(s) and active ingredient(s)) encompass pharmaceuticallyacceptable salts, prodrugs, tautomers, N-oxides, or solvates, e.g.,hydrates, of these materials. It should be understood when reading thisdisclosure that the combination disclosed herein encompasses all theaforementioned variants, as well as any single one thereof orcombination of two or more to less than all such variants.

In certain embodiments, the uveal melanoma can also be metastatic UM.The compounds, methods, and/or combinations described herein can also beused to target metastasis of UM. Particularly, the combination of thedisclosure is suitable for treatment of a patient with UM or metastaticUM, wherein the UM comprises functional p53 or wild-type TP53. Suchprotein or gene status of a cancer is expected to make a patient withsaid cancer even more responsive to the combination of the presentdisclosure. Equally, further improved effect of the combination isexpected in uveal melanoma or metastatic uveal melanoma, includingmetastasis thereof, which is characterized by mutation in either GNAQ orGNA11 genes. In patients harboring both, the functional p53 or wild-typeTP53 and mutation in either GNAQ or GNA11 genes, the clinical responseis expected to be pronounced the most. Therefore, the pharmaceuticalcombination of the present disclosure is best suited for use in thetreatment of a patient with UM or metastatic UM, including UMmetastasis, wherein the UM comprises functional p53 or wild-type TP53and is characterized by mutation in either GNAQ or GNA11 genes.

The term “pharmaceutical combination” as used herein means a productthat results from the use or mixing or combining of more than one activeingredient. It should be understood that pharmaceutical combination asused herein includes both fixed and non-fixed combinations of the activeingredients. The term “fixed combination” means that the activeingredients, e.g., a compound of formula I and one or more combinationpartners, are administered to a patient simultaneously as a singleentity or dosage form. The term in such case refers to a fixed dosecombination in one unit dosage form (e.g., capsule, tablet, or sachet).The terms “non-fixed combination” or a “kit of parts” both mean that theactive ingredients, e.g., a compound of the present disclosure and oneor more combination partners and/or one or more co-agents, areadministered or co-administered to a patient independently as separateentities either simultaneously, concurrently or sequentially with nospecific time limits wherein such administration providestherapeutically effective levels of the two compounds in the body of thepatient, especially where these time intervals allow that thecombination partners show a cooperative, e.g., an additive orsynergistic effect. The term “non-fixed combination” also applies tococktail therapy, e.g., the administration of three or more activeingredients. The term “non-fixed combination” thus defines especiallyadministration, use, composition or formulation in the sense that thecompounds described herein can be dosed independently of each other,i.e., simultaneously or at different time points. It should beunderstood that the term “non-fixed combination” also encompasses theuse of a single agent together with one or more fixed combinationproducts with each independent formulation having distinct amounts ofthe active ingredients contained therein. It should be furtherunderstood that the combination products described herein as well as theterm “non-fixed combinations” encompasses active ingredients (includingthe compounds described herein) where the combination partners areadministered as entirely separate pharmaceutical dosage forms or aspharmaceutical formulations that are also sold independently of eachother. Instructions for the use of the non-fixed combination are or maybe provided in the packaging, e.g., leaflet or the like, or in otherinformation that is provided to physicians and/or medical staff. Theindependent formulations or the parts of the formulation, products, orcompositions, can then be administered simultaneously or chronologicallystaggered, that is the individual parts of the kit of parts can each beadministered at different time points and/or with equal or differenttime intervals for any part of the kit of parts. Particularly, the timeintervals for the dosing are chosen such that the effect on the treateddisease with the combined use of the parts is larger/greater than theeffect obtained by use of only one of the Compounds I-III; thus thecompounds used in pharmaceutical combination described herein arejointly active. The ratio of the total amounts of a compound of formulaI to a compound of formula II or III to be administered as apharmaceutical combination can be varied or adjusted in order to betteraccommodate the needs of a particular patient sub-population to betreated or the needs of the single patient, which can be due, forexample, to age, sex, body weight, etc. of the patients. For example,Compound I can be administered (e.g., orally) at a dose of 100 mg BID,200 mg BID, 300 mg BID, or 400 mg BID in a 28 day cycle while CompoundII is administered (e.g., orally) at a dose of 50 mg, 80 mg, 100 mg or120 mg once weekly for 2 weeks followed by 2 weeks off (Day 1 and Day 8of every 28 day cycle). As yet another example, Compound I can beadministered (e.g., orally) at a lower starting dose (a “run-in” dose),e.g., 50 mg or higher BID (e.g., 50 mg BID, 75 mg BID, 100 mg BID, 200mg BID or 300 mg BID) for a period of time (e.g., first 7 days of every28 day cycle) while compound II is administered at a dose of 50 mg, 80mg, 100 mg, 120 mg once weekly for 2 weeks followed by 2 weeks off,Compound I is then administered (e.g., orally) at a dose higher than therun-in dose, e.g., 75 mg or higher BID (e.g., 200 mg BID, 300 mg BID or400 mg BID) for the rest of days in every 28 day cycle while Compound IIis administered (e.g., orally) at a dose of 50 mg, 80 mg, 100 mg or 120mg once weekly for 2 weeks followed by 2 weeks off (Day 1 and Day 8 ofevery 28 day cycle). In one embodiment, the run-in dose of Compound I is100 mg BID for 7 days and the dose of Compound I following the run-inperiod is 200 mg BID or higher (e.g., 200 mg BID, 300 mg BID or 400 mgBID) while compound II is administered at a dose of 50 mg, 80 mg, 100mg, 120 mg once weekly for 2 weeks followed by 2 weeks off. In anotherembodiment, the run-in dose of Compound I is 200 mg BID for 7 days andthe dose of Compound I in the combination therapy following the run-inperiod is 300 mg BID or higher (e.g., 300 mg BID or 400 mg BID) whilecompound II is administered at a dose of 50 mg, 80 mg, 100 mg, 120 mgonce weekly for 2 weeks followed by 2 weeks off,

The terms “co-administration” or “combined administration” or the likeas utilized herein are meant to encompass the administration of one ormore compounds described herein together with a selected combinationpartner to a single subject in need thereof (e.g., a patient orsubject), and are intended to include treatment regimens in which thecompounds are not necessarily administered by the same route ofadministration and/or at the same time.

The term “pharmaceutical composition” is defined herein to refer to amixture (e.g., a solution or an emulsion) containing at least one activeingredient or therapeutic agent to be administered to a warm-bloodedanimal, e.g., a mammal or human, in order to prevent or treat aparticular disease or condition affecting the warm-blooded animal.

The term “kit of parts” is defined herein to refer to, e.g., combinationpartners (i) and (ii) as defined herein, i.e., (i) at least one proteinkinase C pathway inhibitor (PKCi), e.g., Compound I or apharmaceutically acceptable salt thereof, and (ii) at least one MDM2i,e.g., Compound II or Ill, or a pharmaceutically acceptable salt thereof.The combination partners can be dosed independently or by use ofdifferent fixed combinations with distinguished amounts of thecombination partners (i) and (ii), i.e., simultaneously or at differenttime points. The parts of the kit of parts can then e.g., beadministered simultaneously or chronologically staggered, that is atdifferent time points and with equal or different time intervals for anypart of the kit of parts. The ratio of the total amounts of thecombination partner (i) to the combination partner (ii) to beadministered in the combined preparation can be varied, e.g., in orderto cope with the needs of a patient sub-population to be treated or theneeds of the single patient.

The combination partners in any disclosure embodiment are preferablyformulated or used to be jointly (prophylactically or especiallytherapeutically) active. This means in particular that there is at leastone beneficial effect, e.g., a mutual enhancing of the effect of thecombination partners, in particular a synergism, e.g., a more thanadditive effect, additional advantageous effects (e.g., a furthertherapeutic effect not found for any of the single compounds), less sideeffects, a combined therapeutic effect in a non-effective dosage of oneor both of the combination partners disclosed herein, and verypreferably a clear synergism of the combination partners.

The term “jointly therapeutically active” or “joint therapeutic effect”means that when the therapeutic agents, e.g., the active ingredients,are administered either in a chronologically staggered manner,especially a sequence-specific manner at preferred time intervals, in awarm-blooded animal, especially a human, to be treated, show apreferably synergistic interaction (joint therapeutic effect). Whetherthis is the case can, inter alia, be determined by following the bloodlevels, showing that both compounds are present in the blood of thehuman to be treated at least during certain time intervals.

As used herein, the term “patient” or “subject” refers to an animal.Typically, the animal is a mammal. A patient also refers to for example,primates (e.g., humans), cows, sheep, goats, horses, dogs, cats,rabbits, rats, mice, fish, birds and the like. In certain embodiments,the patient is a primate. In yet other embodiments, the patient is ahuman.

As used herein, the term “carrier” or “pharmaceutically acceptablecarrier” includes any and all solvents, dispersion media, coatings,surfactants, antioxidants, preservatives (e.g., antibacterial agents,antifungal agents), isotonic agents, absorption delaying agents, salts,preservatives, drugs, drug stabilizers, binders, excipients,disintegration agents, lubricants, sweetening agents, flavoring agents,dyes, and the like and combinations thereof, as would be known to thoseskilled in the art (see, for example, Remington's PharmaceuticalSciences, 18th Ed. Mack Printing Company, 1990, pp. 1289-1329). Exceptinsofar as any conventional carrier is incompatible with the activeingredient, its use in the therapeutic or pharmaceutical compositions iscontemplated.

The pharmaceutical combination product according to the disclosure (as afixed combination, or non-fixed combination or as a kit of parts, e.g.,as a combination of a fixed combination and/or individual formulationsfor one or both combination partners or as kit of individualformulations of the combination partners) comprises the combination ofthe present disclosure and one or more pharmaceutically acceptablecarrier materials (carriers, excipients, or the like). Thepharmaceutical combination or the combination partners constituting itcan be formulated for particular routes of administration such as oraladministration, parenteral administration, and rectal administration,etc. In addition, the combination products of the present disclosure canbe made up in a solid form (including without limitation capsules,tablets, pills, granules, powders or suppositories), or in a liquid form(including without limitation solutions, suspensions or emulsions). Thecombination products and/or their combination partners (compounds,active ingredients) can be subjected to conventional pharmaceuticaloperations such as sterilization and/or can contain conventional inertdiluents, lubricating agents, or buffering agents, as well as adjuvants,such as preservatives, stabilizers, wetting agents, emulsifiers andbuffers, etc.

The present disclosure thus pertains to a combination product forsimultaneous or sequential use, such as a combined preparation or apharmaceutical fixed combination, or a combination of such preparationand combination.

In the combination therapies of the disclosure, the compounds usefulaccording to the disclosure may be manufactured and/or formulated by thesame or different manufacturers. Moreover, the combination partners maybe brought together into a combination therapy: (i) prior to release ofthe combination product to physicians (e.g., in the case of a kitcomprising the compound of the disclosure and the other therapeuticagent); (ii) by the physician themselves (or under the guidance of aphysician) shortly before administration; (iii) in the patientthemselves, e.g., during sequential administration of the compound ofthe disclosure and the other therapeutic agent.

The information about the present combination or the use thereof in thetreatment of uveal melanoma as described above and below can be shown ona data carrier, such as for example a product information leaflet, asummary of product characteristics, a brochure, marketing material, aweb page, or when such information is stored or used on a data carriersuch as for example a computer, an USB stick or a CD. Data carriercomprising information about using (i) a PKCi and (ii) an MDM2i isdisclosed. The data carrier, for example in a form of a productinformation leaflet or a label, packaging, brochure or web pageinstruction can be used to instruct to administer (i) a PKCi of formulaI, or a pharmaceutically acceptable salt thereof, and (ii) an MDM2i offormula II or Ill, or a pharmaceutically acceptable salt thereof,simultaneously or sequentially for the treatment of cancer. The datacarrier is particularly useful in the event the two partners of thecombination are not formulated together, and supplied or soldseparately. Each of the partners can be supplied with the data carrier,or even have the data carrier detached or provided separately, thatinforms or instructs about the possibility to use the combinationpartner in a pharmaceutical combination of the present disclosure. Thedata carrier can be used for the same purpose also in fixed combinationsor situations, where both partners are supplied or sold together.

In certain embodiments, any of the above pharmaceutical combination,use, administration, composition, method, product or formulationinvolves further administering one or more other (e.g., third)co-agents.

Thus, the disclosure also relates in a further embodiment to apharmaceutical combination, particularly a pharmaceutical composition ora product comprising a therapeutically effective amount of (i) a PKCiand (ii) an MDM2i, respectively, and at least one third therapeuticallyactive agent (herein referred to as an “additional co-agent”), e.g.,another active ingredient. The additional co-agent is preferablyselected from the group consisting of an anti-cancer agent and ananti-inflammatory agent, particularly is an anti-cancer agent.

The term “a therapeutically effective amount” of a compound (e.g.,chemical entity or biologic agent) of the present disclosure refers toan amount of the compound of the present disclosure that will elicit thebiological or medical response of a subject, for example, reduction orinhibition of an enzyme or a protein activity, or ameliorate symptoms,alleviate conditions, slow or delay disease progression, or prevent adisease, etc. The therapeutically effective dosage of a compound, thepharmaceutical composition, or the combinations thereof, is dependent onthe species of the subject, the body weight, age, sex, and individualcondition, the disorder or disease or the severity thereof beingtreated. A physician, clinician or veterinarian of ordinary skill canreadily determine the effective amount of each of the active ingredientsnecessary to prevent, treat or inhibit the progress of the disorder ordisease.

Frequency of dosage may vary depending on the compound used and theparticular condition to be treated or prevented. In general, the use ofthe minimum dosage that is sufficient to provide effective therapy ispreferred. Patients may generally be monitored for therapeuticeffectiveness using assays suitable for the condition being treated orprevented, which will be familiar to those of ordinary skill in the art.

In certain embodiments, Compound II or III can be generally administeredin unit dosage of about 1-5000 mg of active ingredient(s) for a subjectof about 50-70 kg, or about 1 mg-3 g or about 1-250 mg or about 1-150 mgor about 0.5-100 mg, or about 1-50 mg of active ingredient. The unitdosage may be administered once or repeatedly during the same day, orduring the week. For example, daily dose of between 15 mg/kg and 60mg/kg (e.g., 30 mg/kg for injection or 50 mg/kg for oraladministration), or daily dose of between 100 mg and 1500 mg(particularly between 300 mg and 1000 mg, e.g., 500-800 mg, 500 mg, or800 mg) may be suitable for Compound III. For Compound II, doses betweenabout 10 mg and 1000 mg may be suitable. The weekly dose between about50 mg and 200 mg, particularly between about 75 mg and 150 mg,preferably about 100-120 mg, is expected to be efficacious for CompoundI. Doses of the compounds may require drug holidays. For example, thedosing regimen may include 3 weeks on the drug and 1 week off, 2 weekson and 2 weeks off, or 1 week on and 3 weeks off. The combinationpartners may not be administered according to the same dosing regimen.For example, Compound I can be administered daily (once, twice, ormultiple times daily) while Compounds II or III can be administered onceweekly, every 3 weeks or every 4 weeks. For example, Compounds II can beadministered at, e.g., a dose of 120 mg at day 1 and 120 mg at day 8 (oralternatively on day 6, 7, 9, or 10) in a treatment cycle that lasts 28days (4 weeks).

More examples of MDM2 inhibitors and dosing regimens (e.g., those forCompound II) suitable for the methods, compositions, and combinations ofthe disclosure are described in the Applications U.S. Ser. No.62/422,144 filed Nov. 15, 2016 and 62/479,391 filed on Mar. 31, 2017,the contents of each of which are incorporated herein by reference intheir entireties.

Unless otherwise specified, the weight or dosage referred to herein fora particular compound (e.g., any of Compounds I-III) of the disclosureis the weight or dosage of the compound itself, not that of a salt orprodrug thereof, which can be different to achieve the intendedtherapeutic effect. For example, the weight or dosage of a correspondingsalt of a compound suitable for the methods, compositions, orcombinations disclosed herein may be calculated based on the ratio ofthe molecular weights of the salt and compound itself.

The combination partners (e.g., the individual compounds describedherein) that together form a corresponding pharmaceutical combinationaccording to the disclosure may be mixed to form a fixed pharmaceuticalcomposition or they may be administered separately or at approximatelythe same time (i.e., before, simultaneously with or after the other drugsubstance(s)).

The pharmaceutical compositions that comprise the pharmaceuticalcombination of the disclosure can be tablets or gelatin capsulescomprising the active ingredient together with one or more commonlyknown carriers, e.g., one or more carriers selected from the groupconsisting of

-   -   a) one or more diluents, e.g., lactose, dextrose, sucrose,        mannitol, sorbitol, cellulose and/or glycine;    -   b) one or more lubricants, e.g., silica, talcum, stearic acid,        its magnesium or calcium salt and/or polyethyleneglycol; for        tablets also    -   c) one or more binders, e.g., magnesium aluminum silicate,        starch paste, gelatin, tragacanth, methylcellulose, sodium        carboxymethyl cellulose and/or polyvinylpyrrolidone; if desired    -   d) one or more disintegrants, e.g., starches, agar, alginic acid        or its sodium salt, or effervescent mixtures; and    -   e) one or more absorbents, colorants, flavors and/or sweeteners.

Tablets may be either film coated or enteric coated according to methodsknown in the art.

Suitable compositions for oral administration especially include aneffective amount of one or more or in case of fixed combinationformulations each of the combination partners (active ingredients) inthe form of tablets, lozenges, aqueous or oily suspensions, dispersiblepowders or granules, emulsion, hard or soft capsules, or syrups orelixirs. Compositions intended for oral use are prepared according toany method known in the art for the manufacture of pharmaceuticalcompositions and such compositions can contain one or more agentsselected from the group consisting of sweetening agents, flavoringagents, coloring agents and preserving agents in order to providepharmaceutically elegant and palatable preparations. Tablets may containthe active ingredient(s) in admixture with nontoxic pharmaceuticallyacceptable excipients which are suitable for the manufacture of tablets.These excipients are, for example, inert diluents, such as calciumcarbonate, sodium carbonate, lactose, calcium phosphate or sodiumphosphate; granulating and disintegrating agents, for example, cornstarch, or alginic acid; binding agents, for example, starch, gelatin oracacia; and lubricating agents, for example magnesium stearate, stearicacid or talc. The tablets are uncoated or coated by known techniques todelay disintegration and absorption in the gastrointestinal tract andthereby provide a sustained action over a longer period. For example, atime delay material such as glyceryl monostearate or glyceryl distearatecan be employed. Formulations for oral use can be presented as hardgelatin capsules wherein the active ingredient is mixed with an inertsolid diluent, for example, calcium carbonate, calcium phosphate orkaolin, or as soft gelatin capsules wherein the active ingredient ismixed with water or an oil medium, for example, peanut oil, liquidparaffin or olive oil. Doses of Mdm2 inhibitors used in a compositionmay vary and is dependent for example on the route of administration,gender of a patient, weight, stadium of a disease, etc.

Parenteral compositions and other can be prepared by known methods inthe art.

The use of the articles “a”, “an”, and “the” in both the description andclaims are to be construed to cover both the singular and the plural,unless otherwise indicated herein or clearly contradicted by context.The terms “comprising”, “having”, “being of” as in “being of a chemicalformula”, “including”, and “containing” are to be construed as openterms (i.e., meaning “including but not limited to”) unless otherwisenoted. Additionally whenever “comprising” or another open-ended term isused in an embodiment, it is to be understood that the same embodimentcan be more narrowly claimed using the intermediate term “consistingessentially of” or the closed term “consisting of”.

The term “about”, “approximately”, or “approximate”, when used inconnection with a numerical value, means that a collection or range ofvalues is included. For example, “about X” includes a range of valuesthat are ±20%, ±10%, ±5%, ±2%, ±1%, ±0.5%, ±0.2%, or ±0.1% of X, where Xis a numerical value. In one embodiment, the term “about” refers to arange of values which are 10% more or less than the specified value. Inanother embodiment, the term “about” refers to a range of values whichare 5% more or less than the specified value. In another embodiment, theterm “about” refers to a range of values which are 1% more or less thanthe specified value.

Recitation of ranges of values are merely intended to serve as ashorthand method of referring individually to each separate valuefalling within the range, unless otherwise indicated herein, and eachseparate value is incorporated into the specification as if it wereindividually recited herein. A range used herein, unless otherwisespecified, includes the two limits of the range. For example, the terms“between X and Y” and “range from X to Y, are inclusive of X and Y andthe integers there between. On the other hand, when a series ofindividual values are referred to in the disclosure, any range includingany of the two individual values as the two end points is also conceivedin this disclosure. For example, the expression “a dose of about 100 mg,200 mg, or 400 mg” can also mean “a dose ranging from 100 to 200 mg”, “adose ranging from 200 to 400 mg”, or “a dose ranging from 100 to 400mg”.

The following Examples illustrate the disclosure and provide specificembodiments, however without limiting the scope of the disclosure.

EXAMPLES Example 1: A Phase I Trial of Compound I, a PKC Inhibitor forthe Treatment of Uveal Melanoma

A phase 1, multicenter, open-label, single-arm study of Compound I wasperformed in patients with metastatic uveal melanoma. A Bayesianlogistic regression model, employing the escalation with overdosecontrol principle, was used to guide the dose escalation and to estimatethe maximum tolerated dose (MTD)/recommended dose for expansion (RDE).Safety was described using incidence of adverse events (AEs) by relationand severity of the AE graded according to the Common TerminologyCriteria for AEs (CTCAE, v4.03). Overall response rate was defined asper Response Evaluation Criteria in Solid Tumors (RECIST, v1.1).

Patients eligible for inclusion in this study have to meet all of thefollowing key criteria:

-   -   Metastatic uveal melanoma (histologically or cytologically        confirmed) that is judged to be progressive; patients may be        treatment naive or have had any number of lines of prior        therapy.    -   Measurable disease, defined as at least one lesion that can be        accurately measured in at least one dimension (longest diameter        to be recorded) as >20 mm with conventional techniques or as >10        mm with CT scan.    -   ECOG performance status ≤1

Patients eligible for this study must not meet any of the followingcriteria:

-   -   Malignant disease other than that being treated in this study    -   Symptomatic or untreated leptomeningeal or brain metastases or        spinal cord compression. Treated brain metastases must have been        stable for at least 1 month.    -   Impaired cardiac function or clinically significant cardiac        diseases    -   Patients with abnormal laboratory values as defined by the        protocol    -   Systemic anti-cancer therapy within 2 weeks of the first dose of        study treatment. For cytotoxic agents that have major delayed        toxicity, e.g. mitomycin C and nitrosoureas, and for anti-PD-1        or PD-L1 antibodies 4 weeks is indicated as the washout period.        For patients receiving a CTLA-4 antagonist or vaccine as        anticancer therapy, 6 weeks is indicated as the washout period.        Patients must have recovered or stabilized from toxicities        related to their previous treatment except for alopecia.

As of the data of Jun. 12, 2017, 55 patients received Compound I (i.e.,LXS196) at doses ranging from 100 mg to 1000 mg once daily (QD; 38patients or “pts”) and 200 mg to 400 mg twice daily (BID; 17 pts). Doselimiting toxicities (DLTs) were reported in 7 of 38 pts (QD schedule)and in 1 of 12 pts (BID schedule). The most common DLT was hypotension,manageable with LXS196 interruption and dose reduction. MTDs weredetermined at 500 mg QD and 400 mg BID. Due to better overalltolerability the RDE is 300 mg BID. Most common AEs (all grade, in ≥20%of pts) suspected related to LXS196 in pts across both schedules (n=55)were nausea (58.2%), diarrhea (32.7%), vomiting (25.5%), hypotension(25.5%), fatigue (20%) and asthenia (20%). The majority of GI andconstitutional AEs were grade (gr) ½. Gr ¾ AEs suspected related toLXS196 were reported in 12 pts (21.8%), the most common beinghypotension (10.9%). All other gr ¾ AEs occurred in ≤2 pts (3.6%) each.BID was better tolerated than QD dosing with fewer gr ¾ AEs (5.9% withBID vs 28.9% with QD dosing).

Pharmacokinetic (PK) studies demonstrated rapid absorption of LXS196with T_(max) of ˜1 hr post dose and moderate elimination with consistentterminal T_(1/2) across different doses (˜10 hrs). Exposure at dosesabove 300 mg QD and 200 mg BID is in the efficacious range frompreclinical projections. LXS196 led to reduction of phosphorylatedmyristoylated, alanine-rich C kinase substrate (pMARCKS) andphosphorylated PKC delta (pPKC delta), evident of target engagement inon-treatment tumor biopsies.

In 68 evaluable pts, 6 had confirmed partial responses (PR) per RECIST v1.1 (300 mg QD, 500 mg QD, 200 mg BID, 2 at 300 mg BID, 400 mg BID). Afurther 45 pts had stable disease as best overall response. Preliminarydata suggest encouraging clinical activity of LXS196 as monotherapy withmanageable toxicities in pts with metastatic UM. The single-agent partof the study has completed enrolment.

Example 2: Combination Therapy of Compound I with MDM2i

The anti-tumor activity and tolerability of Compound I (i.e., LXS196)and the MDM2 inhibitor (Compound III or its salt thereof) wasinvestigated in vivo using two patient derived uveal melanoma xenograftmodels, MP46 (GNAQmut) and MP55 (GNA11mut/BAP1 null).

MP46 and MP55 uveal melanoma tumors were established in female SCID miceby subcutaneous implantation of 3×3×3 mm tumor fragments into the flankof each mouse. When tumors reached approximately 100 mm³, mice wererandomized according to tumor volume into treatment groups(n=8-10/group). Test agents were administered orally once (QD) or twicedaily (BID) at the dose levels indicated. Tumor volumes of treatmentgroups are plotted from the time the treatment groups were randomized,and dosing commenced.

Combined oral treatment of LXS196 with Compound III led to increaseddepth of tumor response compared to either single agent in both models.In the MP46 model, Compound III dosed at 100 mg/kg QD achieved 39%treatment/control (T/C), while LXS196 dosed at 120 mg/kg BID achieved63% regression, 35 days post dosing. The combination of LXS196 (dosed at120 mg/kg BID) with Compound III (dosed at 100 mg/kg QD) achievedfurther improvements in anti-tumor response, achieving 97% regression.In the MP55 model, both LXS196 dosed at 120 mg/kg BID and Compound IIIdosed at 100 mg/kg QD achieved 20% T/C. In contrast, the combination ofLXS196 and Compound III led to a 95% regression, 28 days post dosing.Collectively, these data suggest that combined treatment with LXS196 andCompound III may achieve greater and more durable responses in patientswith uveal melanoma.

Efficacy of the combination of Compounds I and II was tested in the UM92.1 cell line. The combination resulted in synergisticanti-proliferative effects (synergy score=2.24). See WO2017/029588,which is incorporated herein by reference.

Other Embodiments

While the invention has been described in conjunction with the detaileddescription thereof, the foregoing description is intended to illustrateand not limit the scope of the invention, which is indicated by thescope of the appended claims. Other aspects, advantages, andmodifications are within the scope of the following claims. All changesthat come within the meaning and range of equivalency of the claims areintended to be embraced therein.

1. A method for treating uveal melanoma comprising administering to a subject in need of thereof 3-amino-N-(3-(4-amino-4-methylpiperidin-1-yl)pyridin-2-yl)-6-(3-(trifluoromethyl)pyridin-2-yl)pyrazine-2-carboxamide (Compound I) or a pharmaceutically acceptable salt thereof at a dose of from about 100 mg to about 1000 mg of Compound I daily.
 2. The method of claim 1, wherein Compound I or a pharmaceutically acceptable salt thereof is administered at a dose of between about 100 mg BID and about 400 mg BID of Compound I.
 3. The method of claim 2, wherein Compound I or a pharmaceutically acceptable salt thereof is administered at a dose of about 50 mg BID, 100 mg BID, 150 mg BID, 200 mg BID, 250 mg BID, 300 mg BID, 350 mg BID, or 400 mg BID.
 4. The method of claim 3, wherein the dose is about 300 mg BID of Compound I.
 5. The method of claim 1, wherein Compound I or a pharmaceutically acceptable salt thereof is administered at a dose of from about 100 mg to about 1000 mg of Compound I once daily.
 6. The method of claim 5, wherein the dose is about 400 mg, 500 mg, 600 mg, 700 mg or about 800 mg once daily.
 7. The method of any one of the preceding claims, wherein the subject suffers from metastatic uveal melanoma.
 8. The method of any one of the preceding claims, wherein the subject is an adult patient aged 18 years or older.
 9. The method of any one of the preceding claims, wherein Compound I or a pharmaceutically acceptable salt thereof is orally administered.
 10. The method of any one of the preceding claims, further comprising administering to the subject one or more other therapeutic agents.
 11. The method of any one of the preceding claims, further comprising administering to the subject a mouse double minute 2 inhibitor (MDM2i).
 12. The method of any one of the preceding claims, further comprising administering to the subject a therapeutically effective amount of mouse double minute 2 inhibitor (MDM2i) selected from (S)-5-(5-chloro-1-methyl-2-oxo-1,2-dihydro-pyridin-3-yl)-6-(4-chloro-phenyl)-2-(2,4-dimethoxy-pyrimidin-5-yl)-1-isopropyl-5,6-dihydro-1H-pyrrolo[3,4-d]imidazol-4-one (Compound II), (S)-1-(4-chloro-phenyl)-7-isopropoxy-6-methoxy-2-(4-{methyl-[4-(4-methyl-3-oxo-piperazin-1-yl)-trans-cyclohexylmethyl]-amino}-phenyl)-1,4-dihydro-2H-isoquinolin-3-one (Compound III), and pharmaceutically acceptable salts thereof.
 13. The method of any one of the preceding claims, further comprising administering to the subject a therapeutically effective amount of (S)-1-(4-chloro-phenyl)-7-isopropoxy-6-methoxy-2-(4-{methyl-[4-(4-methyl-3-oxo-piperazin-1-yl)-trans-cyclohexylmethyl]-amino}-phenyl)-1,4-dihydro-2H-isoquinolin-3-one (Compound III), or a pharmaceutically acceptable salt thereof.
 14. The method of any one of the preceding claims, further comprising administering to the subject a therapeutically effective amount of (S)-5-(5-chloro-1-methyl-2-oxo-1,2-dihydro-pyridin-3-yl)-6-(4-chloro-phenyl)-2-(2,4-dimethoxy-pyrimidin-5-yl)-1-isopropyl-5,6-dihydro-1H-pyrrolo[3,4-d]imidazol-4-one (Compound II), or a pharmaceutically acceptable salt thereof.
 15. The method of any one of the preceding claims, further comprising administering to the subject a therapeutically effective amount of (S)-5-(5-chloro-1-methyl-2-oxo-1,2-dihydro-pyridin-3-yl)-6-(4-chloro-phenyl)-2-(2,4-dimethoxy-pyrimidin-5-yl)-1-isopropyl-5,6-dihydro-1H-pyrrolo[3,4-d]imidazol-4-one (Compound II), or a pharmaceutically acceptable salt thereof once weekly.
 16. The method of any one of claims 12-15, wherein Compound II or III, or a pharmaceutically acceptable salt thereof is orally administered.
 17. The method of claim 15 or 16, wherein Compound II or a pharmaceutically acceptable salt thereof is administered at a dose of from about 50 mg to about 200 mg of Compound II.
 18. The method of any one of claims 15-17, wherein Compound II or a pharmaceutically acceptable salt thereof is administered at a dose of about 50 mg, about 80 mg, about 100 mg, about 120 mg, about 150 mg, or about 200 mg of Compound II.
 19. The method of any one of the preceding claims, wherein the uveal melanoma comprises functional p53 or wild-type TP53.
 20. The method of any one of the preceding claims, wherein the uveal melanoma or metastatic uveal melanoma is characterized by mutation of guanine nucleotide-binding protein G(q) subunit alpha (GNAQ) gene or guanine nucleotide-binding protein G(q) subunit 11 (GNA11) gene. 